Lycopene is a carotenoid substance naturally occurring in tomatoes and many other plant and microbial sources, as a mixture of geometrical isomers. It is an acyclic molecule of formula C40H56, with 11 conjugated and 2 isolated double bonds. (Carotenoids, G. Britton, S. Liaaen-Jensen and H. Pfander, Birkhauser Verlag, Basel, Several volumes, 1995 to 2004). Theoretically there can be many isomers possible. Some of them definitely occur naturally, although it is very difficult to distinguish them from each other by simple spectroscopic methods. For example, the (5-Z) and the all-E isomers have identical UV-Vis spectrum and can be identified only when resolved by RP-HPLC. Eight (mono-Z) isomers were obtained by controlled stereospecific synthesis and six from isomerization mixtures. Four (di-Z) isomers and one (tetra-Z) isomer have also been reported. (Hengartner et al, Helv. Chim. Acta, 75, 1848, 1992). Lycopene from natural sources has been used as a coloring ingredient in foods. It has also been recommended as a useful anti-oxidant. There are no specific reports in the literature on the relative biological activities of the isomers of lycopene. Tomatoes are the major source of lycopene in human nutrition and are known to contain 71 to 90% of all-E-lycopene (all-trans) and 9 to 21% of Z-isomers (cis), mainly the 5-Z-isomer (Zumbrum et al, Helv. Chim. Acta, 68, 1540, 1985), depending on the source, season etc. Surprisingly in humans all-E lycopene accounted for only 12 to 21% whereas the Z-isomers accounted for 79 to 88% in benign or malignant prostate tissue. (Clinton et al., Cancer Epidemiol. Biomarkers Prev., 5, 823, 1996). This means the human body is converting much of the all-E isomer to Z-isomers which may be influencing malignant growth. In beta-carotenes and retinoids the all-E isomers are definitely known to be more active than the Z-isomers. It is thus considered preferable to use all-E lycopene and avoid Z-lycopenes for human consumption. The USP-NF monograph on lycopene is a mixture of all-E lycopene containing up to 23% of 5-Z lycopene, the major isomer in natural lycopene. Synthetic lycopene is better controlled in terms of isomers compared to natural sources. Even so, it is known that the (5-Z)-isomer predominates under certain conditions. A stereospecific synthesis of all-E lycopene suppressing the formation of Z-isomers would be very expensive. A process which enables isomerization of the Z-isomers to all-E isomer is attractive when a synthetic route for lycopene is adopted for its large scale production.
Isomerization of carotenoid compounds is known. Most studies deal with mechanics and mechanisms of photoisomerization and some with enzyme induced isomerizations. (See Dugave and Demange, Chem. Rev., 103, 2481, 2003). Mueller et al. (Pure & Appl. Chem. 69, 2039, 1997) have reviewed the topic and reported that in solution (E/Z)-isomerization of carotenoids is promoted by heat, light, active surfaces and catalytic amounts of acids or iodine. It is believed that the Wittig and Horner condensation steps in the synthesis of carotenoids lead to isomeric mixtures and isomerization could be effected thermally in non-polar solvents. However, the focus in this publication was on preparation of Z-isomers in pure form by isomerization of the all-E lycopene. U.S. Pat. No. 7,126,036 discloses a process of thermal isomerization of lycopene. The process essentially consists of first dissolving the mixture of all-E and Z-isomers in a non-polar solvent dichloromethane followed by addition of methanol, distilling off azeotropically dichloromethane to obtain a suspension in methanol which is then subjected to thermal isomerization by refluxing in methanol or under autogenous pressure raising the temperature to about 95° C. The yields of the enriched all-E isomer or the extent of isomerization did not improve with the autogenous pressure. Although the inventors claimed enrichment of the all-E isomer in mixtures of any proportion of the two isomers, all the examples indicate that the starting mixture consisted of 53% of all-E isomer. The Z-isomer content in the starting mixture is reported as 18%. The inventors achieved enrichment to about 76 to 87% of all-E isomer at the end of the process. The content of the Z-isomer in the enriched mixtures have not been revealed. The extent of isomerization of the Z-isomer to the all-E isomer is thus not clear. Example 7, without thermal isomerization, also achieved enrichment to 75% of all-E isomer in the crystallized sample indicating that solubility and crystallization steps also contributed to a large extent in the process.
We attempted to repeat the thermal isomerization process as described in the U.S. Pat. No. 7,126,036. We used samples containing about 56% of all-E and about 43% of Z-isomers (predominantly 5-Z-isomer) and also containing about 20 to 22% all-E and about 62 to 72% of Z-isomers. Divis product that is routinely obtained from the manufacturing process has this latter composition. The results obtained from these experiments are summarized in the Table 1 below.
TABLE 1Composition ofInput materialReactionoutput materialAll-E (%)Z- (%)time (hrs)All-E (%)Z- (%)Notes56.543.51573.426.5a56.543.53074.825.2a56.543.5Isolated solid74.125.9a22.772.51620.873.1b22.772.54421.272.1b22.772.5Isolated solid24.474.8b28.162.4 648.743.5c128.162.41233.862.1c228.264.21527.572.5d28.264.23030.769.3dNotes:a: 43 g input;b: 25 g input, yield ~22 g;c1: 20 g input, 120° C., 7 bar pressure;c2: 20 g input, 95° C., 1.8 bar pressure;d: 43 kg input (pilot scale).
From the results it can be seen that thermal isomerization of lycopene does not take place when the Z-isomer content is higher than that of all-E isomer. Best result is seen at high temperature and pressure with some enrichment but not satisfying the USP-NF specification of less than 23% of the Z-isomer content. Since lycopene is used as additive to foods and medicaments it was felt desirable to avoid chlorinated hydrocarbon solvents in the final steps of synthesis.
We also attempted to use other solvents in the thermal isomerization experiments. The results are shown in Table 2 below:
TABLE 2Thermal isomerization in different solvents:LycopeneOutput lycopeneall-EZ-(% A)Z- (% A)SolventYieldall-E (% A)(% A)30.457.5TolueneDecomp.——30.457.5Toluene withDecomp.——Iodine (0.01 g)32.654.7IPA80%43.152.932.654.7Pet. ether80%37.253.132.654.7Water84%50.945.030.457.5n-Hexane70%46.249.332.654.7n-Heptane80%36.854.0
It can be seen that the best result was obtained in water as medium in respect of both yield and extent of isomerization. However this was still much below requirement.
There is a need for an improved process of isomerization of Z-isomers present in the synthetic lycopene used as additive in foods or as antioxidant in medicaments. In particular, it was necessary to isomerize mixtures containing low levels of all-E isomer (less than 50%) and high levels of Z-isomers as obtained in commercial manufacture and to avoid halohydrocarbon solvents in the process.